Differential IL-1β secretion by monocyte subsets is regulated by Hsp27 through modulating mRNA stability.

Monocytes play a central role in regulating inflammation in response to infection or injury, and during auto-inflammatory diseases. Human blood contains classical, intermediate and non-classical monocyte subsets that each express characteristic patterns of cell surface CD16 and CD14; each subset also has specific functional properties, but the mechanisms underlying many of their distinctive features are undefined. Of particular interest is how monocyte subsets regulate secretion of the apical pro-inflammatory cytokine IL-1β, which is central to the initiation of immune responses but is also implicated in the pathology of various auto-immune/auto-inflammatory conditions. Here we show that primary human non-classical monocytes, exposed to LPS or LPS + BzATP (3'-O-(4-benzoyl)benzyl-ATP, a P2X7R agonist), produce approx. 80% less IL-1β than intermediate or classical monocytes. Despite their low CD14 expression, LPS-sensing, caspase-1 activation and P2X7R activity were comparable in non-classical monocytes to other subsets: their diminished ability to produce IL-1β instead arose from 50% increased IL-1β mRNA decay rates, mediated by Hsp27. These findings identify the Hsp27 pathway as a novel therapeutic target for the management of conditions featuring dysregulated IL-1β production, and represent an advancement in understanding of both physiological inflammatory responses and the pathogenesis of inflammatory diseases involving monocyte-derived IL-1β

The Second Transmembrane Domain of P2X7 Contributes to Dilated Pore Formation

Activation of the purinergic receptor P2X7 leads to the cellular permeability of low molecular weight cations. To determine which domains of P2X7 are necessary for this permeability, we exchanged either the C-terminus or portions of the second transmembrane domain (TM2) with those in P2X1 or P2X4. Replacement of the C-terminus of P2X7 with either P2X1 or P2X4 prevented surface expression of the chimeric receptor. Similarly, chimeric P2X7 containing TM2 from P2X1 or P2X4 had reduced surface expression and no permeability to cationic dyes. Exchanging the N-terminal 10 residues or C-terminal 14 residues of the P2X7 TM2 with the corresponding region of P2X1 TM2 partially restored surface expression and limited pore permeability. To further probe TM2 structure, we replaced single residues in P2X7 TM2 with those in P2X1 or P2X4. We identified multiple substitutions that drastically changed pore permeability without altering surface expression. Three substitutions (Q332P, Y336T, and Y343L) individually reduced pore formation as indicated by decreased dye uptake and also reduced membrane blebbing in response to ATP exposure. Three others substitutions, V335T, S342G, and S342A each enhanced dye uptake, membrane blebbing and cell death. Our results demonstrate a critical role for the TM2 domain of P2X7 in receptor function, and provide a structural basis for differences between purinergic receptors. © 2013 Sun et al

Plasma membrane cholesterol as a regulator of human and rodent P2X7 receptor activation and sensitization.

P2X7 receptors are nonselective cation channels gated by high extracellular ATP, but with sustained activation, receptor sensitization occurs, whereby the intrinsic pore dilates, making the cell permeable to large organic cations, which eventually leads to cell death. P2X7 receptors associate with cholesterol-rich lipid rafts, but it is unclear how this affects the properties of the receptor channel. Here we show that pore-forming properties of human and rodent P2X7 receptors are sensitive to perturbations of cholesterol levels. Acute depletion of cholesterol with 5 mm methyl-β-cyclodextrin (MCD) caused a substantial increase in the rate of agonist-evoked pore formation, as measured by the uptake of ethidium dye, whereas cholesterol loading inhibited this process. Patch clamp analysis of P2X7 receptor currents carried by Na(+) and N-methyl-D-glucamine (NMDG(+)) showed enhanced activation and current facilitation following cholesterol depletion. This contrasts with the inhibitory effect of methyl-β-cyclodextrin reported for other P2X subtypes. Mutational analysis suggests the involvement of an N-terminal region and a proximal C-terminal region that comprises multiple cholesterol recognition amino acid consensus (CRAC) motifs, in the cholesterol sensitivity of channel gating. These results reveal cholesterol as a negative regulator of P2X7 receptor pore formation, protecting cells from P2X7-mediated cell death.This work was supported by the Biotechnology and Biological Sciences Research Council (BB/F001320/1), the David James Studentship, Department of Pharmacology, University of Cambridge and the Marshall Scholarship.This paper was originally published in The Journal of Biological Chemistry (Robinson LE, Shridar M, Smith P, Murrell-Lagnado RD, The Journal of Biological Chemistry 2014, 289, 46, 31983–31994, doi:10.1074/jbc.M114.574699

Image based machine learning for identification of macrophage subsets

Macrophages play a crucial rule in orchestrating immune responses against pathogens and foreign materials. Macrophages have remarkable plasticity in response to environmental cues and are able to acquire a spectrum of activation status, best exemplified by pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes at the two ends of the spectrum. Characterisation of M1 and M2 subsets is usually carried out by quantification of multiple cell surface markers, transcription factors and cytokine profiles. These approaches are time consuming, require large numbers of cells and are resource intensive. In this study, we used machine learning algorithms to develop a simple and fast imaging-based approach that enables automated identification of different macrophage functional phenotypes using their cell size and morphology. Fluorescent microscopy was used to assess cell morphology of different cell types which were stained for nucleus and actin distribution using DAPI and phalloidin respectively. By only analysing their morphology we were able to identify M1 and M2 phenotypes effectively and could distinguish them from naïve macrophages and monocytes with an average accuracy of 90%. Thus we suggest high-content and automated image analysis can be used for fast phenotyping of functionally diverse cell populations with reasonable accuracy and without the need for using multiple markers

Purinergic signalling and immune cells

This review article provides a historical perspective on the role of purinergic signalling in the regulation of various subsets of immune cells from early discoveries to current understanding. It is now recognised that adenosine 5'-triphosphate (ATP) and other nucleotides are released from cells following stress or injury. They can act on virtually all subsets of immune cells through a spectrum of P2X ligand-gated ion channels and G protein-coupled P2Y receptors. Furthermore, ATP is rapidly degraded into adenosine by ectonucleotidases such as CD39 and CD73, and adenosine exerts additional regulatory effects through its own receptors. The resulting effect ranges from stimulation to tolerance depending on the amount and time courses of nucleotides released, and the balance between ATP and adenosine. This review identifies the various receptors involved in the different subsets of immune cells and their effects on the function of these cells

A community-sourced glossary of open scholarship terms

Supplementary Information: This list of terms represents the ‘Open Scholarship Glossary 1.0’ (available at: https://forrt.org/glossary/. Glossary available under a CC BY NC SA 4.0 license at: https://static-content.springer.com/esm/art%3A10.1038%2Fs41562-021-01269-4/MediaObjects/41562_2021_1269_MOESM1_ESM.pdf).https://static-content.springer.com/esm/art%3A10.1038%2Fs41562-021-01269-4/MediaObjects/41562_2021_1269_MOESM1_ESM.pd

Could It Be Proto Magic? Deceptive Tactics in Nonhuman Animals Resemble Magician’s Misdirection

In the last decade, the study of magic has started to gain the attention of the psychological sciences (Kuhn, 2019; Tompkins, 2019). Psychologists study magic to further our understanding of human cognition including perception, memory, and consciousness. Here, we explore the conscious elements that might make magic unique to humans, such as the experience of wonder and disbelief at the unexpected. We discuss the tactics used by cognitively advanced non-human animals such as apes, corvids, and cephalopods to deceive conspecifics, and consider the parallels between their techniques and the ones used by magicians to make their audience experience the impossible. We also discuss the social dynamics of magic performance, explore the similarities and differences of human and non-human social interactions, and discuss play behaviour in non-human animals. We argue that apes, corvids, and cephalopods might be ideal candidates to start a comparative science of magic, as they appear to naturally exploit analogous blind spots in perception and attention, as well roadblocks in memory, mental time travel and perspective-taking. We highlight the potential of this new and exciting line of research that unlocks alternative avenues for inquiry and investigation. The application of magic to comparative psychology might reveal several interesting psychological constraints across diverse animal minds and offer potential candidates for questions about consciousness

The modern use of ancient theatres related to acoustic and lighting requirements: Stage design guidelines for the Greek theatre of Syracuse

The ancient theatres present today two sides: an architectural one, as archaeological site, and an artistic one, as an entertainment place. This double cultural heritage generates conflicts in the manner of use, so in ancient theatres conservation rules sometimes collide with the intense (and often inappropriate) use during the theatrical summer season.This article presents a research conducted on the Greek theatre of Syracuse related to acoustic and lighting comfort. A parametric study through the prediction software Odeon 10.1 and Relux 2012.1 has been carried out by adding different scenic elements in the theatre to evaluate their influence on acoustic quality and solar radiation. For the acoustic part, the comparison between the conditions with and without the scenery allowed to establish guidelines for scenic design, to preserve and improve the original acoustic apparatus. The computer model was calibrated according to in situ measurements performed in non-occupied conditions. For the lighting part, the natural light analysis during the hours of daytime use of the theatre defined potential elements of discomfort for the viewers, due to glare or high contrasts of luminance between scenery and sky. Through an integrated approach of acoustic and lighting design it has been possible to solve those problems